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Stage #1: With potassium carbonate In water at 20℃; for 0.25 h; Stage #2: for 24 h;
L1 (0.79 g, 3.14 mmol) and K2CO3 (1.04 g, 7.50 mmol) were dissolved in 5.8 mL of water and stirred at room temperature for 15 minutes after which N-(9-fluorenylmethoxycarbonyl) succinimide (Fmoc-OSu, 1.28 g, 3.79 mmol) was added and the mixture was stirred for 24 hours, while the progress of the reaction was monitored by TLC (CHCl3/MeOH/AcOH, 5:1:0.06). The salt was filtered and the filtrate was washed with Et2O, acidified to pH 1 using 3 N HCl, and the desired compound extracted with DCM. After removing the solvent in vacuo, Purification by RP-HPLC, the product-containing fraction was lyophilized to give Compound 11a (1.08 g, yield 73percent). Chemical formula of Compound 11a: C25H1NO8 required [M+H]+=474.2, found [M+H]+=474.9, [M+Na]+=496.9.
With trifluoroacetic acid In dichloromethane for 1.5 h;
Compound 6 was dissolved in DCM (3.5 mL) and TFA (3.5 mL) was added under a nitrogen atmosphere. After 1.5 h of stirring the reaction mixture was concentrated under reduced pressure. Then the excess of TFA was removed by repeated concentration in toluene. DCM/Et2O (100 mL, 1/2, v/v) was added and washed with 1 N HCl. The water layer was extracted with DCM/Et2O (100 mL, 1/2, v/v). The combined organic layers were washed with brine and dried over MgSO4. After filtration the solvent was removed under atmospheric pressure (50°C). The crude oil was purified by silica column chromatography (DCM/MeOH/AcOH, 99/0/1^89/10/1, v/v/v), to give compound 7. Remaining AcOH was removed by dissolving the crude oily product in DCM/Et2O (1/2, v/v) and washing with H2O (3 times) and brine followed by drying over MgSO4. After filtration the solvent was removed under atmospheric pressure (50°C) to give compound 7 as a yellowish oil (0.37 g, 67percent). Rf 0.32 (DCM/MeOH, 89/10/1, v/v).
67%
With trifluoroacetic acid In dichloromethane for 1.5 h;
EXAMPLE 215-λr-(9-FluorenyImethyIoxycarbonyl)-15-aza-3,6,9,12-tetraoxa-pentadecanoate (3) Compound 2 was dissolved in DCM (3.5 mL) and TFA (3.5 mL) was added under a nitrogen atmosphere. After 1.5 h of stirring the reaction mixture was concentrated under reduced pressure. Then the excess of TFA was removed by repeated concentration in toluene. DCM/Et2O (100 mL, 1/2, v/v) was added and the solution was washed with 1 N HCl. The water layer was extracted with DCMZEt2O (100 mL, 1/2, v/v). The combined organic layers were washed with brine and dried over MgSO,*. After filtration the solvent was removed under atmospheric pressure (5O0C). The crude oil was purified by silica gel column chromatography (DCM/MeOH/AcOH, 99/0/1^89/10/1, v/v/v), to give compound 3. Remaining AcOH was removed by dissolving the crude oily product in DCMZEt2O (1/2, v/v) and rinsing with H2O (3x) and brine followed by drying over MgSO4. After filtration the solvent was removed under EPO <DP n="17"/>atmospheric pressure (5O0C) to give compound 3 as a yellowish oil (0.37 g, 67percent). Rf 0.32 (DCM/MeOH/AcOH, 89/10/1, v/v).
With trifluoroacetic acid; In dichloromethane; for 1.5h;
Compound 6 was dissolved in DCM (3.5 mL) and TFA (3.5 mL) was added under a nitrogen atmosphere. After 1.5 h of stirring the reaction mixture was concentrated under reduced pressure. Then the excess of TFA was removed by repeated concentration in toluene. DCM/Et2O (100 mL, 1/2, v/v) was added and washed with 1 N HCl. The water layer was extracted with DCM/Et2O (100 mL, 1/2, v/v). The combined organic layers were washed with brine and dried over MgSO4. After filtration the solvent was removed under atmospheric pressure (50C). The crude oil was purified by silica column chromatography (DCM/MeOH/AcOH, 99/0/1^89/10/1, v/v/v), to give compound 7. Remaining AcOH was removed by dissolving the crude oily product in DCM/Et2O (1/2, v/v) and washing with H2O (3 times) and brine followed by drying over MgSO4. After filtration the solvent was removed under atmospheric pressure (50C) to give compound 7 as a yellowish oil (0.37 g, 67%). Rf 0.32 (DCM/MeOH, 89/10/1, v/v).
67%
With trifluoroacetic acid; In dichloromethane; for 1.5h;
EXAMPLE 215-lambdar-(9-FluorenyImethyIoxycarbonyl)-15-aza-3,6,9,12-tetraoxa-pentadecanoate (3) Compound 2 was dissolved in DCM (3.5 mL) and TFA (3.5 mL) was added under a nitrogen atmosphere. After 1.5 h of stirring the reaction mixture was concentrated under reduced pressure. Then the excess of TFA was removed by repeated concentration in toluene. DCM/Et2O (100 mL, 1/2, v/v) was added and the solution was washed with 1 N HCl. The water layer was extracted with DCMZEt2O (100 mL, 1/2, v/v). The combined organic layers were washed with brine and dried over MgSO,*. After filtration the solvent was removed under atmospheric pressure (5O0C). The crude oil was purified by silica gel column chromatography (DCM/MeOH/AcOH, 99/0/1^89/10/1, v/v/v), to give compound 3. Remaining AcOH was removed by dissolving the crude oily product in DCMZEt2O (1/2, v/v) and rinsing with H2O (3x) and brine followed by drying over MgSO4. After filtration the solvent was removed under EPO <DP n="17"/>atmospheric pressure (5O0C) to give compound 3 as a yellowish oil (0.37 g, 67%). Rf 0.32 (DCM/MeOH/AcOH, 89/10/1, v/v).
L1 (0.79 g, 3.14 mmol) and K2CO3 (1.04 g, 7.50 mmol) were dissolved in 5.8 mL of water and stirred at room temperature for 15 minutes after which N-(9-fluorenylmethoxycarbonyl) succinimide (Fmoc-OSu, 1.28 g, 3.79 mmol) was added and the mixture was stirred for 24 hours, while the progress of the reaction was monitored by TLC (CHCl3/MeOH/AcOH, 5:1:0.06). The salt was filtered and the filtrate was washed with Et2O, acidified to pH 1 using 3 N HCl, and the desired compound extracted with DCM. After removing the solvent in vacuo, Purification by RP-HPLC, the product-containing fraction was lyophilized to give Compound 11a (1.08 g, yield 73%). Chemical formula of Compound 11a: C25H1NO8 required [M+H]+=474.2, found [M+H]+=474.9, [M+Na]+=496.9.
With O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium tetrafluoroborate; N-ethyl-N,N-diisopropylamine; In dichloromethane; N,N-dimethyl-formamide; for 2h;
To a solution of Compound 11a (645.2 mg, 1.36 mmole) in 4 mL DCM and Compound 11b (413 mg, 1.36 mmole) in 2 mL DMF, TBTU (110 mg) and DIPEA (844 mg) were added. The reaction was gone completion after 2 hours. Solvent was removed under a reduced pressure. Inertsil ODS-3 column 30×250 mm; flow rate 32 mL/min; Compound 11c RT, 17 min). Compound 11c was obtained as a white solid (738 mg; yield 76%). Chemical formula of Compound 11c: C39H58N4O11Na. (ESI, positive ion): M/Z: 760.4 (M+1); 782.4 (M+Na). HRMS found 781.4005 for C39H58N4O11Na, err: 1.3 ppm.